Abstract ? Non-alcoholic fatty liver disease (NAFLD) encompasses non-alcoholic fatty liver (NAFL/steatosis) to non-alcoholic steatohepatitis (NASH). Although simple steatosis can be ?benign?, it is an independent risk factor for NASH development. Thiazolidinediones (TZD) are PPAR? agonists used to reduce steatosis in diabetics with NASH. However, the impact of TZD on steatosis is limited, without any resolution of fibrosis. The antisteatotic actions of TZD may be due to reduced insulin resistance in both humans and mouse models. However, TZD-mediated activation of hepatocyte PPAR? may offset the putative antisteatogenic effects of TZD and prevent any reduction in fibrosis. This is based on the following observations: 1) Hepatic PPAR? dramatically increases in NAFLD; 2) TZD exacerbate steatosis in a hepatocyte PPAR?-dependent fashion in mice; and 3) Clinically, the antisteatogenic effect of TZD is limited to the reduction in insulin resistance. To date, the impact of hepatocyte PPAR?-regulated lipid metabolism on NAFLD is poorly understood. Therefore we have knocked-down hepatocyte PPAR? (>99.5%) in adult PPAR?fl/fl mice using an adeno-associated viral vector that express Cre recombinase only in hepatocytes (AAV8-TBGp-Cre). Loss of hepatocyte PPAR? reduced diet-induced steatosis, therefore in SA#1 studies are proposed to determine how loss of hepatocyte PPAR? prevents diet-induced NAFLD. We hypothesize that the reduction in Cd36-mediated FA uptake and/or Mogat1-mediated FA re-esterification, observed after loss of hepatocyte PPAR?, are critical for the reduction in steatosis and subsequent development of NASH. To test this hypothesis, adult PPAR?fl/fl mice will be fed a high fat, high cholesterol, high sucrose diet (HF-HC-HSD) known to induce steatosis (8 weeks of diet) and NASH (27 weeks of diet). Hepatocyte PPAR? will be knocked down, without or with restoration of Cd36 (FA translocase) or Mogat1 (Monoacylglycerol acyltransferase). Hepatic FA uptake, MOGAT activity, FA composition of triacylglycerols (TAG), diacylglycerols (DAG), and monoacylglycerols (MAG)], gene expression and liver and systemic metabolic pathophysiology will be assessed. Although loss of hepatocyte PPAR? reduced steatosis, it led to postprandial dyslipidemia. Therefore in SA#2 studies are proposed to determine the mechanism(s) that promotes postprandial dyslipidemia after loss of hepatocyte PPAR?. We hypothesize that dyslipidemia associated with specific loss of hepatocyte PPAR? is due to enhanced intestinal fat absorption which will be tested by feeding mice a diet containing a nonabsorbable fat or after oral delivery of radio-labeled TAG in the presence of tyloxapol. We also hypothesize that changes in tissue-specific TAG uptake is impaired after loss of hepatocyte PPAR?, which will be tested by assessing liver, heart, muscle and adipose tissue uptake of radio-labeled TAG delivered by oral gavage or iv (to factor out changes in intestinal uptake). The results derived from this project will have an impact in the field because they will define the role of hepatocyte PPAR? and its downstream mechanisms in: 1) the development of diet-induced steatosis and NASH; 2) the dysregulation of metabolic function in NASH; 3) the contribution to hepatic lipid composition to insulin sensitivity and liver disease progression; and 4) the regulation of lipid homeostasis by controlling intestinal lipid absorption and/or tissue-specific postprandial TAG clearance. These results could lead to the development of drugs that block prosteatotic actions of hepatocyte PPAR? that could be used alone or in combination with other therapies to prevent and reverse NASH.